Miquel et al.

Microbial Cell Factories (2015) 14:48

DOI 10.1186/s12934-015-0229-1

REVIEW Open Access

A proposed framework for an appropriate

evaluation scheme for microorganisms as novel
foods with a health claim in Europe
Sylvie Miquel1,2, Martin Beaumont1,2, Rebeca Martín1,2, Philippe Langella1,2, Véronique Braesco3 and Muriel Thomas1,2*

Abstract
This paper concerns the procedure and the scientific approach to obtain market authorization for a microorganism
to be recognized as a novel food with a health claim. Microorganisms that have not been traditionally used during
food production in Europe prior to 1997 are considered as novel foods, which should undergo an in-depth
characterization and safety assessment before being authorized on the European market. If a novel food bacterium
is claimed to provide a beneficial effect on health, these claims must also be investigated before they can be authorized.
Some requirements to obtain novel food certification are shared with those required to obtain a health claim. Although
regulation exists that deals with these issues for foods in general, bacteria in food raise a specific set of questions that
are only minimally addressed in official documentation. We propose a framework and suggest a list of criteria that
should be assessed to obtain marketing authorization and health claim for a bacterium in accordance with European
health policy.
Keywords: Beneficial microbes, Probiotic, Regulation, Safety, Gastro Intestinal (GI) tract

Introduction in theory be considered in itself as a health claim [3]. For

The introduction of high throughput sequencing, ad-
vanced bioinformatics, and specialized in vitro and
in vivo models has improved the understanding of
mechanisms underlying the action of probiotics. Probio-
tics are defined as “live microorganisms which when ad-
ministered in adequate amounts, confer a health benefit
on the host” [1]. For grammatical reasons, this definition
has been recently re-worded by an expert panel of the
International Scientific Association for Probiotics and
Prebiotics (ISAPP) as, “live microorganisms that, when
administrated in adequate amounts, confer a health bene-
fit on the host” [2]. Since February 2013, the European
Commission no longer allows companies to communicate
assumed health benefits of products based solely on pro-
biotic content. Yet, the use of the term “probiotic”, which
is not regulated in Europe, implies that the product has a
beneficial health effect, and thus this designation should
* Correspondence: muriel.thomas@jouy.inra.fr
1
Commensal and Probiotics-Host Interactions Laboratory, UMR1319 Micalis,
INRA, AgroParisTech, Domaine de Vilvert, 78350 Jouy en Josas, France
2
AgroParisTech, UMR 1319 MICALIS, F-78350 Jouy-en-Josas, France
Full list of author information is available at the end of the article
instance, the Food Safety Authority of Ireland clearly indi-
cates on its website that the “term probiotic is considered
to be a health claim”.
Emerging clinical evidence suggest that beneficial bac-
teria positively influence a wide range of human health
issues, especially digestive health [4-7]. Today, most mi-
croorganisms marketed as “probiotics” or beneficial bac-
teria by the food industry belong to the genera
Lactobacillus and Bifidobacterium [8]. However, these
genera are sub-dominant in the intestinal microbiota in
adults. This observation, in association with rapidly
expanding knowledge of the human microbiome, sug-
gests that a large panel of potential new candidates can
be isolated from the dominant members of our adult
microbiota. The real challenge for translational projects
between scientists and industrial partners will be the
introduction of new generations of beneficial strains be-
cause there is currently a large gap between the bench
and the market. It is indeed difficult for all stakeholders,
including academic and industrial partners, to agree
unanimously on a system of regulation, which may need
to be adjusted on a case-by-case basis.

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Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article,
unless otherwise stated.
Miquel et al. Microbial Cell Factories (2015) 14:48
This review focuses on foods and food ingredients
consisting of or including live bacteria and does not
cover genetically modified microorganisms, which raise
other issues. Bacteria with no history of documented safe
use in Europe prior to 1997 are classified as novel foods.
Requirements that must be fulfilled for such bacteria
to be allowed on the market include the accurate
characterization of the strain and a solid demonstra-
tion of its safety. If a novel food confers a beneficial ef-
fect on health, it will become a novel food associated
with a health claim after approval by the EFSA and
authorization from the European Commission. How-
ever, it remains difficult to define experiments that can
be judged as reliable, valuable, and pertinent to sup-
port the application of novel bacteria. Here, we
summarize criteria that are commonly suggested by
authorities or in the scientific literature for the
characterization of novel bacteria and the assessment
of their safety and efficacy. We also present our point
of view about the biological relevance and regulatory
significance of these criteria and of the experimental
methods often proposed to meet them.
What is a novel food according to the current European
food regulation?
Many microorganisms that are used as food ingredients
or food additives have a long history of safe use in food
fermentations [8]. However, microorganisms that were
not traditionally used in food production in Europe be-
fore 1997 are classified as a novel food class 2.2 (see
Table 1 and Figure 1). The market and the legislation of
novel foods are regulated by the UE 97/618/EC recommen-
dation and regulation No 258/97 (Table 1 and Figure 1).
The launch a Novel Food in Europe is authorized if the
product has been thoroughly characterized and its safety
has been proven (Figure 1).
For a novel food to be allowed on the market, those
submitting the application are required to do the
following:
# Characterize the microorganism and its metabolites
# Characterize how the process of production may
modify the microorganism
# Give the history of production and consumption of
the microorganism
# Anticipate the intake/extent of the use and the
consumption of the novel food
# Provide the nutritional composition of the novel food
# Assess the safety of the microorganism
# Give toxicological information
An example of an application for a novel food is the
public version of an application for the use of Clostridium
butyricum (CBM588), (which was isolated from a soil
Page 2 of 11
Table 1 Statutory text, official documents of the EFSA
Novel Food
Recommendation Related to the scientific aspects and the
97/618/EC presentation of information necessary to support
applications to place novel foods and novel food
ingredients on the market
Regulation Related to novel foods and novel food ingredients of
N° 258/97 the European Parliament and of the European Council
Novel Food Complex Novel Food from non-Genetically Modified
Class 2 sources according to Categories of novel foods and
novel food ingredients identified in Regulation (EC)
No 258/97. Intact plants, animals and microorganisms
used as foods as well as food components (e.g.
complex carbohydrates, fats, proteins or those
substances collectively described as dietary fiber)
are included. Two sub-classes can be identified:
2.1 the source of the NF has a history of use in
food in the Community.
2.2 the source of the NF has no history of use in
food in the Community.
Health claim
Regulation Related to nutrition and health claims and establishes
N° 1924/2006 rules governing the Community authorization of
health claims should only be authorized in the
Community after a scientific assessment of the highest
possible standard, to be carried out by the EFSA.
sample), as a novel food supplement in the European
Union (EU) (Miya-Pro; Public version – Clostridium
butyricum 588 novel food application).
What is a novel food with a health claim according to the
current European food regulation?
A novel food is not required to have any beneficial nutri-
tional or health-related effects upon consumption. How-
ever, a health claim would highlight the benefits of the
novel food for the consumer and may help its commer-
cial success (Figure 2). The process for substantiating a
health claim for a novel food is the same as for a con-
ventional food. Beneficial nutritional or health claims
may be communicated to the consumer only after
authorization from the European Commission, which re-
quires a favorable opinion from the EFSA according to
EU regulation 1924/2006 (see Table 1 and Figure 1). As
shown in Figure 2, some requirements to obtain novel
food certification are the same as those required to cer-
tify a health claim (e.g. strain characterization).
Overall, most bacteria that will be used in foods for
human consumption in the near future will need to
comply with two different regulations (EC 258/97 and
EC 1924/2006), which largely involve scientific require-
ments. Successful market applications will require vari-
ous skills allying the academic and industrial worlds, to
address the numerous regulatory, economic, and scien-
tific challenges.
Miquel et al. Microbial Cell Factories (2015) 14:48 Page 3 of 11

For future foods and food ingredients consisting of or isolated from bacteria

Could you prove the

consumption within the Yes No
EU before 15 May 1997

Is it a Novel Food ? No Yes

Do you need to fulfil the UE

No Yes
258/97 ?

For future health claims

Do you need to fulfil

Yes Yes
The UE 1924/2006?
Figure 1 Statutory text.

How can current regulation be improved to ensure the
correct characterization of future novel-food microorganisms
with a health claim?
Characterization of the strain of microorganism is the
first requirement both to apply for novel food market-
ing authorization and to submit a health claim request
(Figure 2). Functional effects are only related to the
particular strain and cannot be extended to the rest of
the species to which the strain belongs [9,10]. However,
the only approved health claim is for yoghurt cultures,
which are not defined at the strain level [11]. Thus, if
there is a reasonable scientific basis to justify a health
benefit for an entire species, then the health claim need
not be limited to one strain. Moreover, the ISAPP re-
cently proposed the creation of a general category of
probiotics defined at the species level and associated
with core benefits [2]. It is also important to consider
characterizing the strain in the various environments
Novel foods Health claims
EU 258/97 EU 1924/2006
Compulsory Optional
Consumption
history Health effect
Characterization evidence
Safety
Bringing to market Sales success
Figure 2 Novel food or health claim?
that it may encounter from the start of the production
process until the final consumption of the product.
The EFSA recommends that both the species (DNA-
DNA hybridization or 16S rRNA gene sequence analysis)
and strain (genetic typing molecular methods) be identi-
fied [12]. Indeed, it is essential to identify clearly the spe-
cies and strain of bacteria based on phenotypic and
genotypic data [13] from various methods considered as
reliable references in the literature (Table 2) [14]. Al-
though all methods listed in Table 2 are relevant, some
may be replaced by whole genome sequencing technolo-
gies, which are undergoing rapid development. In
addition to characterizing the strain, the sequence may
reveal the pathogenic potential of the strain and identify
virulence genes; thus, this information may be relevant
for the toxicological assessment required for a novel
food. Additionally, genomic data are useful to search for
functional relationships between genetic elements and
the mechanisms underlying probiotic action [15]. Puta-
tive proteins and metabolic pathways can also be pre-
dicted from genomic sequences and these predictions
can be confirmed by metabolomic and transcriptomic
profiling with high-throughput techniques. In light of
the development of many “omic” techniques, it would be
interesting to establish a strain-specific map of genomic
and functional interactions. In the near future, it is prob-
able that most bacteria that will be used for the first time
in foods for human consumption will have to be entirely
sequenced.
Clearly, whole genome sequencing is the gold standard
for characterization; however, we can only make sense of
these data if we consider a strain in its physiological
context and take into account its overall biology. Life
cycle, industrial processes, storage history and ingestion
may affect the phenotypic properties of bacteria and
the potential activities of a particular strain [16]. It
could be worthwhile to describe the properties of
microorganisms during different physiological states
Miquel et al. Microbial Cell Factories (2015) 14:48
Table 2 Criteria and methods often proposed to characterize a strain (required for both novel food and health claim
regulations)
Taxonomic Biological Methods
level assignment
Species Phenotypic GRAM staining
Morphologic description
Analytical profile identification (API)
Genotypic 16S rRNA sequencing [15,33]
MLST (Multi Locus Sequence Typing)
Strain Metabolic Analytical profile identification (API)
Genotypic Genome sequencing [15]: Identification of virulence genes
Pulsed field gel electrophoresis (PFGE) [20]: evaluation of
genetic stability [58]
(latency, multiplication, and stationary phases) but also
in the context of the various micro-environments that
the product could encounter. However, the exact na-
ture of such tests cannot be generalized and specific
tests are needed for each microorganism used. For in-
stance, the yoghurt bacteria, Streptococcus thermophilus
and Lactobacillus delbrueckii ssp. bulgaricus present
different growth rates and metabolic activities depend-
ing on the culture media (milk, presence of lactose or
rich media) [17-19]. Although the environment may in-
fluence the metabolic activities and/or functions of mi-
croorganisms the inverse is also true: microorganisms may
affect the physico-chemical conditions of their environ-
ment. Moreover, strains could evolve and functional muta-
tions could appear. Thus, pulsed field gel electrophoresis
(PFGE) may be informative because a high rate of muta-
tions may lead to the appearance of new characteristics
[20]. This is mainly a typing technology, yielding limited
data on functionality. Most (functionally-important) SNPs
are actually not detected by rare cutting restriction en-
zyme analysis. Thus, novel-food bacteria should also be
highly characterized in the various environments that they
will encounter from production to consumption.
The final product of the food matrix in which the
microorganism is present should also be precisely
characterized:
– to assure the absence of microbial contaminants [21];
– to define the composition of the food because it may
affect strain growth;
– to assure proper labeling of the food with regard to
macro-nutrients, calories and allergens, especially
for particular consumers (e.g. diabetics) [22];
– to define clearly the conditions of use (the target
population, the storage conditions and any
precautions about the time and regularity of intake)
and to support its claimed effect [23].
Page 4 of 11
Propositions for improvement
The physiology of the bacterium in environments
from production to consumption should be
extensively characterized
Full genome sequencing should be systematically
provided
Concerning intake, it is strongly recommended that
the final products contain an adequate amount of live
bacteria to provide a health benefit [24,25]. A daily in-
take of at least 108–109 viable cells, which may be
achieved by the daily consumption of at least 100 g of
final product, has been suggested as the minimum intake
to provide an effect [24].
How should the safety of microorganisms and their
metabolites be assessed? Some suggestions to improve
current regulatory requirements
In the EU, the a priori safety of some microorganisms is
accepted if they benefit from Qualified Presumption of
Safety (QPS) status [26].
Most of these microorganisms are Gram positive, non-
sporulating or lactic acid bacteria [24]. Lactic acid bacteria,
mainly Bifidobacterium, Lactobacillus and Streptococcus
are found in many food products and are not dominant in
the intestinal microbiota in adults [8,22,27-29]. If the
microorganism is not recognized as QPS, a complete as-
sessment of its safety must be carried out according to
regulatory requirements. Some adverse side effects that
must be monitored are the production of host-deleterious
metabolites, systemic infection, inappropriate immune re-
sponses, antibiotic (AB) resistance and gene transfer [30].
However, this list is not exhaustive and there is currently
no official document summarizing these criteria. Some re-
quirements are mentioned in the scientific literature
[21,31,32] and in the Guidelines for the Evaluation of Pro-
biotics in Food published in 2002 by the FAO/WHO
working group [13]. However, the biological relevance of
these requirements remains a subject of debate. We have
ordered and evaluated the criteria that are often proposed
to demonstrate the safety of microorganisms into five clas-
ses: survival and/or viability along the Gastro Intestinal
(GI) tract, effect on intestinal homeostasis, adhesion,
metabolic activities, and remote effects (Table 3).
 Miquel et al. Microbial Cell Factories (2015) 14:48
Table 3 Common criteria generally considered as essential for the safety of NF/probiotic products (required for both novel food and health claim regulations)
What How Why Comments and Propositions for improvement
Survival in GI tract Resistance to intestinal stress In vitro Growth curves/Detection in feces Resistance to GI tract conditions may Not valuable for all beneficial effects
conditions after consumption favor the beneficial effects
Development of new protectors/encapsulators
Bile salt deconjugation High-performance liquid chromatography Large amounts of deconjugated bile Evaluation of property in vitro has poor relevance;
salts may have undesirable effects assessment of bile salt deconjugation in vivo
on the human host
Mass spectrometry
Preservation of the Microbiota Perturbation of commensal In vitro production of bacteriocins or Bacteriocins and AB may perturb Development of growth inhibitory references
homeostasis of gut consortium antibiotics (AB) microbiota. with major commensal bacteria
barrier components
AB may interact with a patient’s
treatment
Antibiotics (AB) resistance In silico*prediction and in vitro antibiogram AB resistance may be transmitted Development of in vivo assessment (animal model)
between bacteria indicating the microbiota homeostasis (composition
Minimal inhibitory concentration test (MIC) and activities) after probiotic consumption
Minimal bactericidal concentration test If plasmids are detected: the presence/absence of
(CMB) genes encoding the most common resistance
determinants should be characterized
Presence of plasmids In silico*prediction or DNA extraction Plasmids favor the transmission of Requirement to up-date the antibiotic list
followed by analysis by gel electrophoresis antibiotic resistance
Mucus Mucus degradation Mucin degradation test (agarose gel or Excessive mucus degradation may The capacity to degrade mucus seems to be a poor
liquid culture) lead to intestinal barrier weakening criterion to estimate the protective or deleterious
effect of bacteria on the intestinal barrier
Adhesion and Intestinal/Mucosal adhesion Test bacterial strain adhesion to epithelial Mucosal adhesion may interfere with The intestinal/mucosal adhesion capacity can be
translocation risk cell line pathogenic microorganisms, stimulate either a beneficial or a deleterious criterion
beneficial cellular processes, or favor
bacterial translocation
Intestinal mucosa degradation Gelatinase activity assay Mucosal degradation may weaken It could be useful to evaluate in vivo
the intestinal barrier translocation capacities
Hemolytic activity Blood agar culture Hemolysis damages red blood cells
Metabolic activities D-Lactate production Colorimetric assay D-Lactate accumulation in blood The production of D-lactate should be compared
leads to acidosis with the amount produced by usual strains
(like in yoghurt)
Toxin production Protocol recommended by the European Toxic molecules Establishment of threshold values relevant in
scientific committee of animal nutrition. humans
Biogenic amine production Colorimetric assay Immune responses such as allergic
responses
Remote effects Platelet aggregation Aggregation test Risk of thrombosis Development of ex vivo protocols (explants,
organoids)
Genotoxicity Toxicity testing on animals models (chronic Risk of cancer

Page 5 of 11
and subchronic tests)
Allergenicity Allergic response
(*: if the complete genome is available).
Miquel et al. Microbial Cell Factories (2015) 14:48
Survival and /or viability in the intestinal tract
It is commonly accepted that bacteria must be able to
survive in at least the upper part of the GI tract to have
a beneficial effect on the host (except bacteria used to
improve microbial composition in the oral cavity). An
intestinal strain would have to resist extreme GI tract
conditions: an intestinal pH gradient (from 4.0 in stom-
ach to 7.0 in the lower part of intestine), bile salts, and
pancreatic secretions [33]. All these variables can be
monitored in vitro by growth curves, although such as-
says do not totally reflect in vivo conditions (Table 3).
The amount of bacteria in vivo can also be determined
by PCR or by isolating and culturing them from stools a
few days after ingestion [34]. However, it is debatable
whether a bacterium must survive and proliferate along
the GI tract to have an effect. For example, polysacchar-
ide A of Bacteroides fragilis protects animals from colitis
induced by Helicobacter hepaticus [35] and the peptido-
glycan of the Lactobacillus salivarius strain Ls33 also
protects against experimentally-induced colitis [36].
Thus, proliferation of the bacterium at the target site is
not absolutely required for its effect and a daily con-
sumption of product may suffice. Nonetheless, survival
in the GI tract needs to be examined in each particular
case, and it would be informative to establish a sensitiv-
ity profile of each strain towards pH, bile salts, and pan-
creatic secretions. Innovative technologies, based for
example on encapsulation, are currently being developed
to control better the viability of microbes during storage,
processing, and in the GI tract [37].
The deconjugation of bile salts is a property that al-
lows bacteria to survive in the GI tract and is implicated
in lowering circulating cholesterol levels [38]; however,
in excess, this process may be deleterious for the host
[39]. Thus, it has been proposed that the bile salt decon-
jugating activity of probiotics should be evaluated [40],
because it could influence bacterial survival and host
health. Evaluation of this property (Table 3) in vitro may
provide little relevant information for the situation
in vivo because of its complexity, raising the need, as a
second step, to perform experiments with an animal
model (mono-associated or with a complete microbiota).
It is yet not completely clear whether bile salt deconju-
gating activity is a desirable trait in a novel food/pro-
biotic bacterium. In our opinion, the capacity to
deconjugate bile salts is informative when choosing a
strain and should therefore be considered.
Preservation of intestinal homeostasis by maintaining the
integrity of two barrier components of the gut: microbiota
and mucus
A key safety requirement is that the use of a new novel
food should not perturb the population commensal or-
ganisms. It may be useful to examine whether novel food
Page 6 of 11
strains inhibit the growth of commensal bacteria, to as-
sess whether and how homeostasis is maintained. This
approach requires the assessment in vitro of the produc-
tion of bacteriocins or antibiotics (AB). It would be use-
ful to test in vitro if probiotics have bactericidal effects
on the main groups of commensal bacteria such as Bac-
teroidetes and Clostridii, although this is not required by
authorities and is rarely proposed in the literature. Then,
the effect of the probiotic consumption on microbiota
composition could be assessed with in vivo experiments
measuring the global metabolic activities of microbiota
(such as the production short-chain fatty acids).
Novel food strains should not be able to transmit anti-
biotic resistance genes to bacteria in their environment
to avoid the acquisition and spread of multiple antibiotic
resistance [33]. The transmission potential of resistance
genes depends on their genetic support (plasmids or
chromosome). Unlike resistance genes carried by plas-
mids, those carried by chromosomes have a much lower
risk of transfer. Thus, it is important to consider the
genomic location of an antibiotic resistance gene when
testing a strain for antibiotic resistance [32]. A list of an-
tibiotics which should be tested to assess antibiotic re-
sistance has been proposed by the EFSA [41]. This list
should be updated according to current treatments and
the target population (Table 3).
It is also widely reported that bacteria are beneficial if
they do not degrade mucus. The rationale is that degrad-
ing mucus weakens the intestinal barrier and conse-
quently destabilizes the protective function of the
epithelium [42]. However, some commensal bacteria use
mucins, the major constituents of mucus, as an energy
source and can stimulate host mucus production [43].
Thus, such adaptive cross-talk does not necessarily im-
pair host defense mechanisms. Moreover, mucin degrad-
ation tests (agarose gel or liquid culture) often use
synthetic mucins from pig gastric mucus. This experi-
mental setting does not accurately represent the typical
features of human intestinal mucus, and moreover, the
diversity of available energetic substrates in vivo is much
higher than in vitro. Thus, bacteria that degrade mucus
in vitro (where mucus is the only energy source in cul-
ture medium) may not necessarily use this substrate
in vivo. In our opinion, the capacity to degrade mucus,
as assessed by in vitro assays, is not a relevant criterion
to estimate the protective or deleterious effect of bac-
teria on the intestinal barrier (Table 3). For instance, ad-
ministration of living Akkermansia muciniphila, which is
a mucin-degrading commensal, reverses dietary induced
metabolic disorders [44].
Adhesion properties and translocation risk
Adhesion to the mucosal layer is commonly mentioned
as a factor favoring durable implantation and a highly
Miquel et al. Microbial Cell Factories (2015) 14:48
effective probiotic [13]. This suggests that transient per-
sistence and/or long-term colonization is associated with
beneficial mechanisms such as interfering with the
growth of pathogenic or potentially pathogenic microor-
ganisms in the body or stimulating other potentially
beneficial cellular processes [45]. In our opinion, this as-
sumption is questionable, because proximity to the in-
testinal mucosa and a long transit time in the gut are
not sufficient to maximize the beneficial effects of a
strain (Table 3). Indeed, although bacteria are rapidly
eliminated, they may act transitorily [46]. For instance,
the immune system of germ-free mice can be stimulated
by temporary bacterial colonization [47]. These results
imply that some microorganisms do not have to colonize
permanently the microbiota to affect host responses
[46]. In addition, in vitro tests have many limits, because
adhesion properties and mucus production depend on
the cell line under study [48]. We believe that although
adhesion is an important characteristic of a strain, it
should not be a criterion to estimate the potential effect
within the gut. However, in vitro tests that assess adhe-
sion also reveal the cytotoxicity of the bacterium on tar-
geted cells. It may be relevant to estimate bacterial
translocation because oral treatments containing a high
dose of probiotic may be deleterious, especially in im-
munodeficient patients [49].
Metabolic activities: threshold and physiological state matter
It is well known that some bacterial metabolites have
deleterious effects; however, the current recommenda-
tions provided by authorities are unclear and no max-
imal threshold for these metabolites has been proposed
(Table 3). For instance, bacteria-induced D-lactate pro-
duction can be viewed as harmful because the accumula-
tion of this metabolite in blood may be neurotoxic and
may lead to acidosis [50]. Individuals the most at risk
are those with short-bowel syndrome (which results
from resection of the small intestine) because of the ac-
cumulation D-lactate in their feces [51]. In healthy
adults, lactate is not detectable in fecal samples because
lactobacilli (main producers of D-lactate) represent a
minor group in the microbiota and lactate is degraded
by other major bacterial groups [52]. Some strains of L.
delbrueckii subsp. bulgaricus that are widely used in yog-
hurt fermentation produce D-Lactate [17]. The subse-
quent risk of acidosis related to D-lactate after the
ingestion of yoghurt is not a health threat in healthy in-
dividuals [53]. Nearly all lactic acid bacteria, even those
widely used for food applications, produce the D-lactate
isoform. Moreover, many commensal strains present in
the lower GI tract consume D-lactate resulting in cross-
feeding that may help to explain the reported butyro-
genic effect of certain dietary substrates [52]. Thus, the
production of D-lactate is not a risk for healthy people.
Page 7 of 11
It remains informative to determine the level of D-
lactate production for each microorganism, but the
production of D-lactate should not be considered sys-
tematically as a metabolic disadvantage for future
health-related strains. It could be interesting to com-
pare the amount of D-lactate produced by the consid-
ered strains with that produced by Lactobacilli currently
present in yoghurt.
Lack of toxin or biogenic amine production is essential
to ensure strain safety but experimental settings and
threshold values are still poorly defined for human appli-
cations. The EFSA recommends a protocol originally
established for animal nutrition and limited to the use of
Bacillus species [54]. This may be too specific to be ex-
trapolated to all strains used in human nutrition or
health. A relevant alternative strategy may be the use of
commensal bacteria as references (Table 3).
Two other tests are frequently mentioned: strain
hemolytic activity and platelet aggregation ability. Both
are typical features of pathogenic bacteria (Table 3).
However, these deleterious effects may only happen if
the ingested bacteria end up in the blood. This is an un-
likely situation which requires bacterial translocation
across a weakened intestinal barrier, which would also
favor the translocation of all commensal bacteria. These
tests however provide important information about
strain pathogenicity [55-57].
Remote effects: toxicology testing
It is necessary to evaluate the toxicological profile, in-
cluding genotoxicity, to establish the safety of a new
product [58]. The harmful effects of a particular sub-
stance or microorganism are examined, evaluated, and
interpreted by testing it on animal models (Table 3).
These results are then extrapolated to determine the
quantity that will produce similar effects in humans.
However, current ethics statements encourage a reduc-
tion in the numbers of animals that are used in these
tests. Thus, it may be useful to develop new techniques
based on viable ex vivo tissue explants or organoids
(from experimental rodents or humans) that could be
co-incubated with bacteria of interest or their respective
metabolites [59,60]. In our opinion, ex vivo intestinal
cultures co-incubated with bacteria or their culture
supernatant are promising approaches that should be
encouraged to evaluate immune responses (cytokine re-
lease in the supernatants and the expression level of
immune receptors), epithelium homeostasis (prolifera-
tion, differentiation and/or apoptosis) and toxicity
markers (e.g. genotoxicity) of new products.
Overall, although in vitro testing can be very inform-
ative as a first step, the properties of candidate probio-
tics should always also be assessed in vivo because they
are strongly influenced by the intestinal ecosystem, the
Miquel et al. Microbial Cell Factories (2015) 14:48
complexity of which cannot be entirely reproduced
in vitro (biochemical properties, interaction with com-
mensal microorganisms or with the host and substrate
availability).
Are health claims based on a cause-effect relationship
between the intake of a microorganism and a benefit to
human health?
Clearly, the safety of a microorganism is essential for the
authorization of a novel food. However, convincing evi-
dence demonstrating the beneficial effects of a particular
microorganism should also be submitted for scientific
evaluation to the Nutrition panel of the EFSA. Numer-
ous general and specific guidelines (depending on the
health claim) have been published by the EFSA and
should be consulted and followed [23,61-63]. Health ef-
fects are demonstrated in the same way regardless of
the nature of the active component of the novel food
(microorganism, nutrient, or any other ingredient).
Successful applications preferably involve studies of
high methodological and statistical quality examining a
well-defined health benefit that can be unambiguously
evaluated through changes in a recognized biomarker
in humans.
It is particularly difficult to define the beneficial effects
after consumption of a bacterium. The ISAPP considers
that beneficial microbes that support a healthy digestive
tract and a healthy immune system are associated with
common general benefits to human health [2]. Indeed,
many of the most promising benefits of bacteria involve
their effect on the gut, and their interactions with either
the gut microbiota or the cells of the intestinal mucosa,
especially epithelial and immune cells. These aspects
have been extensively studied, and although knowledge
is rapidly expanding, the complexity of these interactions
is also becoming increasingly apparent. This has pre-
vented clear conclusions from being made about the ap-
propriate markers to examine the effects of a food or a
microorganism on human health. As a result, a gap ex-
ists between productive research and the effective imple-
mentation of these findings in the life of the consumer.
This is a good illustration of the challenges of transla-
tional research. In addition to research on probiotics
and their effects, there is also a strong and sustainable
need to document markers and to assess the clinical sig-
nificance of probiotics on human health. Hopefully, pro-
biotics have many potential activities and although
efforts should focus on the microbiota and immune sys-
tem, other effects that are clearly beneficial to humans,
can be more readily assessed. A non-exhaustive list of
potential benefits includes reduction in cholesterol
levels, decrease of intestinal pain, favored intestinal tran-
sit, and reinforcement of the intestinal epithelium.
Page 8 of 11
The intestinal microbiota: a potential source of novel
foods with a health claim
The intestinal microbiota, including commensal bacteria
and probiotics, plays a fundamental role in the develop-
ment and maintenance of intestinal homeostasis by par-
ticipating in the immune and digestive functions of the
GI tract [33]. This homeostasis is crucial for the host
health and if disrupted, it may lead to an inappropriate
reaction of the immune and digestive systems.
Newly discovered intestinal bacteria may be used for
the development of new novel foods containing microor-
ganisms with a health claim [2]. Over the past 20 years, in-
tensive research has led to the in depth characterization of
intestinal communities, particularly as a function of age,
health status, geographic location, nutritional habits, med-
ical care, and genetic predisposition of the host [64]. It is
now widely accepted that intestinal commensal microor-
ganisms participate in the physiology and the health of
their host through metabolic, protective, and trophic func-
tions [65]. Host physiology, gut maturation, innate and ac-
quired immune responses and metabolism are largely
influenced by the metabolic properties of the microbiota
[66-69]. The activity and composition of the microbiota
are modulated by external factors, such as diet, making it
a highly “malleable” tissue in humans [70]. It was recently
proposed that Faecalibacterium prausnitzii, which is a
major constituent of the intestinal human microbiota,
may have prophylactic or therapeutic applications in
human health [71,72]. In particular, F. prausnitzii is de-
pleted in many intestinal disorders and displays benefi-
cial anti-inflammatory effects on host, suggesting that
it may be used to counterbalance the dysbiosis linked
to certain diseases [73,74]. Thus, the characterization
of several microbial communities from our environ-
ment (particularly those of our microbiota) may help to
identify new bacterial species with beneficial effects on
human health. Interestingly, the recent description of
the intestinal metagenome (i.e. all genomes of the bac-
terial populations present in the intestine), by sequen-
cing strategies has confirmed that the microbial
richness of the human gut microbiome correlates with
metabolic markers [75]. Therefore, besides being a
huge reservoir of unexploited commensal bacteria, our
microbiota also has metabolic capacities that are poten-
tially beneficial to human health. We can speculate that
this large panel of commensal organisms contains new
promising candidates that may be very efficient in the
digestive tract because they will be reintroduced into
their endogenous ecological niche. However, although
the GI tract is their natural niche, most potentially
commensal probiotic bacteria have never been used in
food products; thus, manufacturers would have to apply
for marketing authorization according to the regulation
of novel foods.
Miquel et al. Microbial Cell Factories (2015) 14:48
Conclusion
We propose a framework that will help academic and in-
dustrial communities to explore the potential of bacteria
as novel foods with health claims in accordance with
European health and nutrition policy. New research on
the human microbiome will facilitate the development
of mechanistically-driven probiotics. This approach
clearly offers a new strategy that may benefit the health
of the general population and that of patients with lim-
ited therapeutic options; for example, it may provide an
alternative to long-term antibiotic use. Further insight
into the precise mechanisms of action of new probiotic
strains may lead to the development of second gener-
ation probiotics with characterized beneficial effects.
Until now, yoghurt is the only probiotic food with a
health claim [11]. It remains unclear whether these regu-
lations limit or boost creativity and innovation [76]. It is
in the interest of stakeholders that this translational sub-
ject, at the cross roads of scientific, industrial, and clin-
ical research, is clarified by appropriate regulations [77].
These regulations clearly indicate, to companies as well
as to risk assessors and managers, that claims should be
based only on very strong scientific evidence. Moreover,
it seems necessary to have some flexibility regarding in-
dividual studies depending on the particular microorgan-
ism involved, the claim area, the target population, and
the condition of use. In this “point of view” paper, we
have discussed some of the tests proposed for the devel-
opment of intestinal probiotics, bearing in mind that in-
novative strategies should be encouraged.
Competing interests
The author(s) declare that they have no competing interests.
Authors’ contributions
SM, VB, MT raised the necessity to review Novel Food regulation and
conceived of the work. SM, MB, RM, MT drafted the manuscript, illustrations
and tables. MT coordinated the work, PL funded financial supports for SM
and RM. PL, VB and MT provided the final version. All authors read and
approved the final manuscript.
Acknowledgements
We thank Claire Cherbuy, Muriel Mercier-Bonin, Françoise Rul, and Joanna
Radziwiłł-Bieńkowska for fruitful discussions. SM and RM received funding
from the FPARIS collaborative project, which was selected and supported by
the Vitagora Competitive Cluster and funded by the French FUI (Fond
Unique Interministériel; FUI: n°F1010012D), the FEDER (Fonds Européen de
Développement Régional; Bourgogne: 34606), the Burgundy Region, the
Conseil Général 21, and the Grand Dijon.
Author details
1
Commensal and Probiotics-Host Interactions Laboratory, UMR1319 Micalis,
INRA, AgroParisTech, Domaine de Vilvert, 78350 Jouy en Josas, France.
2
AgroParisTech, UMR 1319 MICALIS, F-78350 Jouy-en-Josas, France.
3
VAB-nutrition, Clermont-Ferrand, France.
Received: 14 November 2014 Accepted: 17 March 2015
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